To construct basic fibroblast growth factor (bFGF) eukaryotic expression vector and to evaluate the possibility of bFGF gene therapy in orthopedic disease, the pCD-rbFGF recombinant plasmid was constructed by cloning rat basic fibroblast growth factor (bFGF) cDNA into an eukaryotic expression vector, pcDNA3. Rat osteoblasts were transfected with pCD-rbFGF plasmid by lopofectin mediated gene transfer, the transient expression was detected by streptavidin-biotin-enzyme complex (SABC) method. It was observed that the expression of rat bFGF gene was detected 72 h after transfected distinctly. Basic fibroblast growth factor gene therapy is a method of potential for a wide array of orthopedic diseases.
Cells, Cultured ; DNA, Complementary/genetics ; Escherichia coli/genetics ; Eukaryotic Cells/metabolism ; Fibroblast Growth Factor 2/biosynthesis ; Fibroblast Growth Factor 2/*genetics ; Gene Transfer Techniques ; Osteoblasts/cytology ; Osteoblasts/*metabolism ; Plasmids ; Recombinant Proteins/biosynthesis ; Recombinant Proteins/genetics ; *Transfection ; Transformation, Genetic

Adult ; Aged ; Female ; Fibroblast Growth Factor 1 ; genetics ; Fibroblast Growth Factor 2 ; genetics ; Humans ; Middle Aged ; Neoplasm Staging ; Neoplasms, Glandular and Epithelial ; metabolism ; pathology ; Ovarian Neoplasms ; metabolism ; pathology ; RNA, Messenger ; analysis ; Receptor Protein-Tyrosine Kinases ; genetics ; Receptor, Fibroblast Growth Factor, Type 1 ; Receptors, Fibroblast Growth Factor ; genetics

This study was aimed to investigate the influence and significance of celecoxib (specific inhibitor of cyclooxygenase-2) on mRNA expressions of vascular endothelial growth factor (VEGF), basic fibroblast growth factor (b-FGF), transforming growth factor β (TGF-β) in acute leukemia cells. The expressions of VEGF, b-FGF, TGF-β mRNA were measured by RT-PCR in acute leukemia cells treated with celecoxib (80 µmol/L, for 48 h) or with PBS. The results showed that the obvious expressions of VEGF, b-FGF, TGF-β mRNA were observed in acute leukemia cells. By using Pearson correlation analysis, there was positive correlation between VEGF mRNA and b-FGF mRNA expressions (r = 0.559, P = 0.001), and negative correlation between VEGF and TGF-β mRNA expressions (r = -0.4, P = 0.029). Expression levels of VEGF, b-FGF, TGF-β mRNA in experimental group were lower than that in control group (P < 0.01). It is concluded that COX-2 inhibitor celecoxib can inhabit vascular endothelial growth through down-regulating the mRNA expression of VEGF, b-FGF and TGF-β in acute leukemia cells. COX-2 inhibitor may offer supplemental effect for treating acute leukemia.
Adult ; Celecoxib ; Cyclooxygenase 2 Inhibitors ; pharmacology ; Female ; Fibroblast Growth Factor 2 ; metabolism ; Humans ; Leukemia ; drug therapy ; metabolism ; Male ; Pyrazoles ; pharmacology ; RNA, Messenger ; genetics ; Signal Transduction ; Sulfonamides ; pharmacology ; Transforming Growth Factor beta ; metabolism ; Vascular Endothelial Growth Factor A ; metabolism

OBJECTIVETo study the expressions of basic fibroblast growth factor (bFGF) and its receptor (bFGFR) in bone marrow of mice with acute radiation injury, and to evaluate the effect of Ligustrazine (Lt) on them.

METHODSFifty-six Kunming mice of clean grade were randomly divided into 3 groups, the normal group, the control group and the Lt group. Mice in the latter two groups were once homogeneously systemic irradiated with 6.0 Gy of 60Co, with the absorption dose rate of 0.56 Gy/min, then treated with saline (0.2 ml/mice) or Lt (2 mg/mice) respectively, twice a day through gastrogavage for successive 13 days. Mice were sacrificed in batch on the 3rd, 7th and 14th day by cervical dislocation to collect the bilateral femoral bone marrow for preparing bone marrow mono-nuclear cell (BMMNC) suspension. The bFGFR expression on surface of BMMNC was determined by flow cytometry; and the bFGF expression level in one side of femoral bone marrow tissue was detected by immunohistochemistry with SABC-AP assay.

RESULTSThe bFGF expression in bone marrow of mice on the 3rd, 7th and 14th day after acute radiation injury all were significantly lower than that of the normal mice (P < 0.05 or P < 0.01). The expressions of bFGF and bFGFR in the Lt group detected were significantly higher than that in the control group detected at the corresponding time points (P < 0.05 or P < 0.01).

CONCLUSIONBy way of enhancing bFGF expression in bone marrow and bFGFR expression on surface of BMMNC to accelerate the repairing of homopoietic micro-environment in bone marrow might be one of the mechanisms of Lt in promoting hemopoietic function reconstitution after acute radiation injury.

Animals ; Bone Marrow Cells ; metabolism ; Female ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Hematopoiesis ; drug effects ; Male ; Mice ; Pyrazines ; pharmacology ; Radiation Injuries, Experimental ; metabolism ; Random Allocation ; Receptors, Fibroblast Growth Factor ; biosynthesis ; genetics

OBJECTIVETo investigate approach and possibility of transferring basic fibroblast growth factor (bFGF) gene into rabbit bone marrow stromal cells (BMSCs).

METHODSThe eukaryotic expression vectors harboring bFGF cDNA were constructed and transfected into rabbit BMSCs mediated by liposome. The transcription and expression of bFGF gene in the transfected BMSCs were detected by means of morphological observation, immunohistochemistry, enzyme-linked immunosorbent assay (ELISA) and RT-PCR. The changes in the biological characteristics of the transfected MSCs were also observed.

RESULTSStable overexpression of bFGF protein was detected in the transfected BMSCs, which showed differentiation towards chondrocyte lineage.

CONCLUSIONStable expression of bFGF gene in transfected BMSCs can induce cell differentiation into chondrocyte lineage.

Animals ; Bone Marrow Cells ; cytology ; metabolism ; Female ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Gene Expression ; Genetic Vectors ; genetics ; Male ; Rabbits ; Stromal Cells ; cytology ; metabolism ; Transfection

OBJECTIVETo study the effect of adenovirus-mediated basic fibroblast growth factor(FGF-2) gene transfer in vivo on oligodendrocyte cell numbers throughout ventrolateral white matter following spinal cord injury in rats.

METHODSThirty-two adult female Sprague Dawley rats were injured with the Infinite Horizon Impactor, and then were randomly assigned to four groups: FGF-2-Adts high-titer group (1.27x10(7) pfu/rat), FGF-2-Adts intermediate-titre group (6.37x10(6) pfu/rat), FGF-2-Adts low-titer group (3.18 x 10(6) pfu/rat), and green fluorescent protein (GFP)-Adts group (5.9x10(7) pfu/rat). The transgenic expression in vivo was detected with fluorescence microscopy. The locomotor function of the hindlimbs of rats was evaluated using Rivlin plate. Slides mounted with tissue sections were processed for immunohistochemical detection and quantification of oligodendrocytes (CC1(+)) in the ventral lateral funiculi (VLF) of injured spinal cords.

RESULTSOne week after spinal cord injury, GFP showed that many cells had expressed objective gene in vivo and the angles of the occlusal plane of rats in FGF-2 groups were significantly higher than in GFP-Adts group. Also, there was a significant difference among the FGF-2-Adts treatment groups for the volume of gray matter sparing. However, there were no significant differences for total white matter sparing. Stereological quantification of total CC1(+) cell numbers in the spared VLF showed a significant reduction in numbers with GFP controls compared to all other groups 4 weeks after injury. In contrast, the FGF-2 Adts intermediate-titer group had significantly more CC1(+) cells when compared to both the FGF-2-Adts high- and low-titer groups.

CONCLUSIONAdenovirus-mediated FGF-2 gene transfer can promote the functional recovery of the injured spinal cord by enhancing the proliferation and/or differentiation of oligodendrocytes.

Adenoviridae ; genetics ; Animals ; Disease Models, Animal ; Female ; Fibroblast Growth Factor 2 ; genetics ; metabolism ; Genetic Therapy ; Oligodendroglia ; pathology ; Rats ; Rats, Sprague-Dawley ; Spinal Cord Injuries ; metabolism ; physiopathology ; therapy ; Transfection

Cell Line ; Epidermal Growth Factor ; biosynthesis ; genetics ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Hepatocytes ; metabolism ; Humans ; Interleukin-10 ; pharmacology ; Platelet-Derived Growth Factor ; pharmacology ; Proto-Oncogene Proteins c-sis ; RNA, Messenger ; biosynthesis ; genetics

OBJECTIVE: To explore the effects of exogenous transforming growth factor-beta 1 (TGFbeta1) on peripheral nerve regeneration after the peripheral nerve injury and if TGFbeta1 regulates the expression of basic fibroblast growth factor (bFGF) in the anterior horn motoneurons of spinal cord during regeneration. METHODS: Forty-eight rats were crushed on the right sciatic nerve and then randomly divided into 2 groups: TGFbeta1 group and NS group. In TGFbeta1 group, TGFbeta1 50 microL (0.1 microg/mL) was injected into the proximal nerve near to the crushed nerve and after the operation the injured leg was injected with equal TGFbeta1 whereas the NS was replaced in the NS group. The rats of each group survived for 3, 7, 14 and 21 days after the lesion. The bFGF expression in the anterior horn motoneurons of spinal cord was detected by immunohistochemistry (IHC). Semi-thin section and Fast Blue retrograde tracing were also performed with the rats surviving for 21 days to observe the regeneration of distal end in the injured right sciatic nerve. RESULTS: The number of bFGF immunoreactive positive motoneurons in TGFbeta1 group was obviously higher than that of the NS group (P < 0.05). In the distal sciatic nerve of the rats treated with TGFbeta1, the number and diameter of regenerating myelinated axons and the thickness of myelinated sheath were more than those of the NS group (P < 0.05). The number of motoneurons in spinal cord and neurons in dorsol root ganglia (DRG) labelled with Fast Blue in the NS group was obviously lower than in the TGFbeta1 group (P < 0.01). CONCLUSION Exogenous TGFbeta1 plays an important role in promoting the peripheral nerve regeneration; TGFbeta1 up-regulates the bFGF expression in the anterior horn motoneurons of spinal cord during the peripheral nerve regeneration.
Animals ; Female ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Male ; Motor Neurons ; metabolism ; Nerve Regeneration ; drug effects ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Sciatic Nerve ; injuries ; metabolism ; physiology ; Spinal Cord ; metabolism ; Transforming Growth Factor beta ; pharmacology ; Transforming Growth Factor beta1

To investigate the changes in the expression of basic fibroblast growth factor (bFGF) and transforming growth factor beta 2 (TGFbeta2) in glomus cell grafts of carotid body in the rat model of 6-hydroxydopamine-induced Parkinson disease, immunohistochemical staining of bFGF and TGFbeta2 in the sections of striate body was done on the 2nd, 4th and 12th week after transplantation. The results showed that on the 2nd week after transplantation, bFGF and TGFbeta2 were not detectable in the glumous cell grafts. On the 4th week after graft, bFGF and TGFbeta2 immunoreactivity was increased within the grafts and at the graft-host interface but was restricted only to astrocytes. In the striatum surrounding the graft, bFGF was expressed persistently, while TGFbeta2 showed transient expression. It was suggested that the transient expression of TGFbeta2 was likely due more to the trauma imposed by the graft procedure than to an intrinsic. The deficiency in astrocytic bFGF early after graft may be responsible for the poor survival of grafted glomus cells of carotid body.
Animals ; Carotid Body ; cytology ; transplantation ; Female ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Hydroxydopamines ; Parkinson Disease ; etiology ; metabolism ; surgery ; Rats ; Transforming Growth Factor beta ; biosynthesis ; genetics ; Transforming Growth Factor beta2 ; Transplantation, Homologous

OBJECTIVETo elucidate the effects of exogenous basic fibroblast growth factor (bFGF) on biological characteristics of rat osteoblasts cultured in vitro.

METHODSThe osteoblasts isolated from a Sprague-Dawley rat and cultured in vitro were treated with different concentrations of bFGF (5-50 ng/ml) respectively. At 24 hours after treatment, the proliferating cell nuclear antigen was measured with immunocytochemistry, alkaline phosphatase (ALP) activity was determined and the expression of transforming growth factor beta 1 (TGF-beta(1)) was detected to observe the effects of bFGF on growth and differentiation of osteoblasts.

RESULTSbFGF (5-50 ng/ml) could obviously promote the growth of osteoblasts. The intracellular expression of TGF-beta(1) mRNA increased significantly, but the intracellular ALP content decreased.

CONCLUSIONSbFGF can obviously stimulate the proliferation of osteoblasts and promote the synthesis of TGF-beta(1), but cannot promote the differentiation of osteoblasts.

Alkaline Phosphatase ; metabolism ; Animals ; Cells, Cultured ; Fibroblast Growth Factor 2 ; pharmacology ; Osteoblasts ; drug effects ; metabolism ; Proliferating Cell Nuclear Antigen ; analysis ; RNA, Messenger ; analysis ; Rats ; Rats, Sprague-Dawley ; Transforming Growth Factor beta ; genetics ; Transforming Growth Factor beta1